JPWO2007111283A1 - Glycohemoglobin concentration measuring method and concentration measuring apparatus - Google Patents

Abstract

The present invention relates to a method and apparatus for measuring glycated hemoglobin concentration by an optical technique. As the measurement wavelength, a wavelength at which the molecular extinction coefficient of oxyhemoglobin and the molecular extinction coefficient of deoxyhemoglobin match or substantially match is adopted. Preferably, the measurement wavelength is set to a wavelength range of 417 to 421 nm. In the present invention, glycohemoglobin concentration is measured using a sample prepared from red blood cells in blood, for example, using column chromatography.

Description

  The present invention relates to a method for measuring the concentration of glycohemoglobin contained in a sample such as blood.

  When a biological component is separated and analyzed using a biological sample such as blood, a high performance liquid chromatography device (HPLC device) using high performance liquid chromatography (HPLC) is widely used (see, for example, Patent Document 1).

  As shown in FIG. 10, the general HPLC apparatus 9 prepares a sample containing a biological component in the sample preparation unit 90 and then introduces the sample into the analysis column 91. It is comprised so that it may adsorb to. When glucohemoglobin is measured using whole blood as a sample, a biological sample in a state in which the hemolyzed blood is diluted is introduced into the analysis column 91 after hemolyzing red blood cells collected from the whole blood. On the other hand, the biological component adsorbed on the filler is eluted by supplying the eluent from the eluent bottle 93 to the analysis column 91 by the liquid feed pump 92. The eluent from the analysis column 91 is introduced into the photometric mechanism 94, and the photometric mechanism 94 continuously measures the absorbance of the eluent to analyze the biological components.

  As shown in FIG. 11, the photometric mechanism 94 emits light from the light source 97 while the eluent flows through the flow path 96 of the photometric cell 95, and the transmitted light at that time is received by the light receiving unit 98. It is. The wavelength of light received by the light receiving unit 98 is selected by the interference filter 99, while an output level signal corresponding to the amount of light received is output from the light receiving unit 98.

  Further, the HPLC apparatus 9 calculates the total hemoglobin amount based on the chromatogram, which is a change in absorbance with time, and calculates the glycohemoglobin concentration as a proportion of the total hemoglobin amount occupied by glycohemoglobin.

  However, since the amount of dissolved gases such as oxygen in the eluent differs depending on the temperature of the eluent, the biological components were analyzed when the temperature outside the device (environment temperature) fluctuated or when the environment temperature was different. In this case, the dissolved gas state (dissolution amount) in the eluent is different. For this reason, when the dissolved oxygen concentration in the eluent varies with a change in environmental temperature, the ratio of oxyhemoglobin and deoxyhemoglobin in hemoglobin varies. Further, even in the biological sample introduced into the analysis column 91, the ratio between oxyhemoglobin and deoxyhemoglobin in hemoglobin may vary.

  On the other hand, as the biological sample to be introduced into the analysis column 91, the hemolyzed blood is diluted and a relatively large amount of oxygen is used. Therefore, in the photometric mechanism 94, the maximum absorption wavelength of oxyhemoglobin is 415 nm. Adopted as measurement wavelength. For this reason, in an environment where the change in environmental temperature is large, the ratio of oxyhemoglobin and deoxyhemoglobin is different, so accurate measurement becomes difficult when they are measured at the same wavelength.

JP-A-7-120447

  An object of the present invention is to enable appropriate measurement of the concentration of clicohemoglobin even when the ratio between oxyhemoglobin and deoxyhemoglobin is different.

  In the first aspect of the present invention, a glycohemoglobin concentration is measured by an optical method, wherein the measurement wavelength is set to a wavelength at which molecular extinction coefficients of oxyhemoglobin and deoxyhemoglobin match or substantially match. A method for measuring hemoglobin concentration is provided.

  Preferably, the measurement wavelength is set to 417 to 427 nm. More preferably, the measurement wavelength is set to 419 to 425 nm. Of course, the measurement wavelength is not limited to the above range, and other wavelength ranges known as wavelengths where the molecular extinction coefficients of oxyhemoglobin and deoxyhemoglobin match or approximately match, for example, 520-526 nm or 583-589 nm. It may be set.

  The present invention can be applied to the case of measuring glycohemoglobin concentration using column chromatography or using a sample prepared from red blood cells in blood.

  According to a second aspect of the present invention, there is provided a glycated hemoglobin concentration measuring apparatus comprising a photometric mechanism that irradiates a sample with light from a light source and receives light traveling from the sample at that time in a light receiving unit, The photometric mechanism is provided with a glycated hemoglobin concentration measuring apparatus configured such that light having a wavelength at which molecular extinction coefficients of oxyhemoglobin and deoxyhemoglobin match or substantially match can be received by the light receiving unit.

  The photometric mechanism is configured such that, for example, light of 417 to 427 nm can be received by the light receiving unit. Preferably, the photometric mechanism is configured so that light of 419 to 425 nm can be received by the light receiving unit. The photometry mechanism may also be configured so that light of 520 to 526 nm or 583 to 589 nm can be received by the light receiving unit.

  The glycated hemoglobin concentration measuring apparatus of the present invention further includes a separation unit for separating glycated hemoglobin using, for example, column chromatography.

  The glycated hemoglobin concentration measuring apparatus of the present invention may further include a sample preparation mechanism for preparing a sample from red blood cells in blood.

It is a schematic block diagram which shows the HPLC apparatus which is an example of the glycohemoglobin measuring apparatus of this invention. It is sectional drawing for demonstrating the photometry mechanism in the HPLC apparatus shown in FIG. It is a graph which shows the wavelength dependence of the molecular extinction coefficient of oxyhemoglobin and deoxyhemoglobin. FIG. 2 is a block diagram showing a main part of the HPLC apparatus shown in FIG. It is a flowchart for demonstrating operation | movement of the HPLC apparatus shown in FIG. It is a flowchart for demonstrating the density | concentration calculation process in the calculating circuit in the HPLC apparatus shown in FIG. It is an example of the chromatogram obtained in the HPLC apparatus shown in FIG. 2 is a graph showing the relationship between environmental temperature and glycated hemoglobin concentration in Example 1. It is a graph which shows the relationship between the environmental temperature in Example 2, and a glycohemoglobin density | concentration. It is a schematic block diagram which shows the HPLC apparatus which is an example of the conventional glycohemoglobin measuring apparatus. It is sectional drawing for demonstrating the photometry mechanism in the HPLC apparatus shown in FIG.

Explanation of symbols

1 Glycohemoglobin concentration measuring device 3 Sample adjustment unit 5 Photometric mechanism 53B Light receiving element (light receiving part)

  Hereinafter, the present invention will be specifically described with reference to the drawings.

  The HPLC apparatus X shown in FIG. 1 corresponds to an example of the glycated hemoglobin concentration measuring apparatus of the present invention, and is configured to measure the glycated hemoglobin concentration using whole blood. The HPLC apparatus X includes a plurality of eluent bottles 10, 11, 12 (three in the drawing), a deaeration device 2, a sample preparation unit 3, an analysis unit 4, a photometric mechanism 5, and an arithmetic circuit 6.

  Each eluent bottle 10, 11, 12 holds an eluent to be supplied to an analysis column 40 described later. As eluents, for example, buffers having different pH and salt concentrations are used.

  The degassing device 2 is for removing dissolved gas from the eluent before supplying the eluent to the analysis unit 4 (analysis column 40). The degassing device 10 is connected to the eluent bottle 10 via the pipes 70A, 70B, and 70C. 11 and 12 are connected to the manifold 41 of the analysis unit 4 via pipes 71A, 71B and 71C.

  As shown in FIG. 1, the sample preparation unit 3 is for preparing a sample to be introduced into the analysis column 40 from the blood cell component of the blood sample 14 collected from the blood collection tube 13. The sample preparation unit 3 includes a sampling nozzle 30, a preparation liquid tank 31, and a dilution tank 32.

  The sampling nozzle 30 is for collecting various liquids including the blood sample 14 of the blood collection tube 13 and is capable of sucking and discharging the liquid and movable in the vertical and horizontal directions. Yes. The operation of the sampling nozzle 30 is controlled by a control means (not shown).

  The preparation liquid tank 31 holds a preparation liquid for preparing a sample for introduction to be introduced into the analysis column 40 based on the blood sample 14. In the preparation liquid tank 31, a hemolysis agent for hemolyzing red blood cells, a dilution liquid for diluting the hemolysis, and the like are held as preparation liquids.

  The dilution tank 32 is for providing a place for lysing red blood cells in the blood sample 14 and preparing a sample for introduction by diluting the hemolyzed blood. The dilution tank 32 is connected to an injection valve 43 in the analysis unit 4 to be described later via a pipe 72 so that an introduction sample prepared in the dilution tank 32 can be introduced into the analysis column 40 via the injection valve 43. It is configured.

  The analysis unit 4 controls the adsorption / elution of biological components to the packing material of the analysis column 40 and supplies various biological components to the photometric mechanism 5, and the temperature is controlled by a temperature control mechanism (not shown). . The set temperature in the analysis unit 4 is about 40 ° C., for example. The analytical column 40 holds a filler for selectively adsorbing hemoglobin in the sample. As the filler, for example, a methacrylic acid ester copolymer is used.

  The analysis unit 4 has a manifold 41, a liquid feed pump 42, and an injection valve 43 in addition to the analysis column 40.

  The manifold 41 is for selectively supplying the eluent from the specific eluent bottles 10, 11, 12 of the plurality of eluent bottles 10, 11, 12 to the injection valve 43. The manifold 41 is connected to the deaeration device 2 via pipes 71A, 71B, 71C, and is connected to the injection valve 43 via a pipe 73.

  The liquid feed pump 42 is for applying power for moving the eluent to the analysis column 40 via the injection valve 43, and is provided in the middle of the pipe 73. The liquid feed pump 42 is operated so that the flow rate of the eluent becomes 1.0 to 2.0 ml / min, for example.

  The injection valve 43 collects a certain amount of introduction sample and enables the introduction sample to be introduced into the analysis column 40, and includes a plurality of introduction ports and discharge ports (not shown). An injection loop 44 is connected to the injection valve 43. The injection loop 44 is capable of holding a fixed amount (for example, several μL) of liquid, and the injection loop 44 communicates with the dilution tank 32 by appropriately switching the injection valve 43 so that the injection loop 44 is connected to the dilution tank 32. The injection sample is supplied to the analysis column 40 via the pipe 74, and the introduction sample is introduced from the injection loop 44 into the analysis column 40, or the injection loop 44 is not shown in the figure. The state in which the cleaning liquid is supplied from the cleaning tank can be selected. As such an injection valve 43, for example, a six-way valve can be used.

  As shown in FIG. 2, the photometric mechanism 5 is for optically detecting hemoglobin contained in the eluent from the analysis column 40. The photometric cell 50, the light source 51, the beam splitter 52, and the light receiving for measurement. A system 53 and a reference light receiving system 54 are provided.

  The photometric cell 50 is for defining a photometric area. The photometric cell 50 has an introduction channel 50A, a photometry channel 50B, and a discharge channel 50C, and these channels 50A, 50B, and 50C communicate with each other in series. The introduction flow path 50A is for introducing the eluent from the analysis column 40 (see FIG. 1) into the photometric flow path 50B, and is connected to the analysis column 40 via a pipe 75. The photometric flow path 50B circulates the eluent to be measured and provides a field for photometric measurement of the eluent, and is formed in a straight line. The photometric flow path 50B is open at both ends, and both ends are closed by the transparent cover 55. The discharge channel 50C is for discharging the eluent from the photometric channel 50B, and is connected to the waste liquid tank 15 via a pipe 76 (see FIG. 1).

  The light source 51 is for irradiating light to the eluent flowing through the photometric channel 50B. The light source 51 is arranged in a state of facing the end surface 50Ba (transparent cover 55) of the photometric flow channel 50B so that the optical axis L passes through the center of the photometric flow channel 50B. As the light source 51, a light source that can emit light having a measurement wavelength that matches or substantially matches the molecular extinction coefficients of oxyhemoglobin and deoxyhemoglobin and light having a reference wavelength is used. Examples of the measurement wavelength include a wavelength range of 417 to 426 nm, but the measurement wavelength may be 520 to 526 nm or 583 to 589 nm. As the reference wavelength, for example, 500 nm can be adopted. For example, a halogen lamp can be used as the light source 51 capable of emitting light of the measurement wavelength and the reference wavelength in the previous wavelength range. Of course, as the light source 51, other than the halogen lamp, for example, one provided with one or a plurality of LED elements can be used.

  The beam splitter 52 divides the light emitted from the light source 51 and transmitted through the photometric flow path 50B so as to enter the light receiving system 53 for measurement and the light receiving system 54 for reference. Above, it is arranged in a state inclined by 45 degrees. As the beam splitter 52, various known ones such as a half mirror can be used.

  The light receiving system 53 for measurement selectively receives light having a target wavelength among the light transmitted through the beam splitter 52 and is disposed on the optical axis L. The measurement light receiving system 53 includes an interference filter 53A and a light receiving element 53B for receiving light transmitted through the interference filter 53A. As the light receiving element 53B, a photodiode can be used.

  The interference filter 53A selectively transmits light having a target measurement wavelength. Here, in the HPLC apparatus X, the measurement wavelength is set to 417 to 426 nm, preferably 419 to 423 nm. This is due to the following reason. First, as shown in FIG. 3, the molecular extinction coefficient of oxyhemoglobin and the molecular extinction coefficient of deoxyhemoglobin match when the wavelength is approximately 421 nm. Therefore, if the measurement wavelength is 421 nm or a wavelength close thereto, This is because it is considered that the hemoglobin concentration can be measured in the hemoglobin contained in the eluent from the analysis column 40 without being affected by the ratio of oxyhemoglobin to deoxyhemoglobin or the variation thereof. Second, as will be apparent from the examples described later, since the result that the measurement wavelength is 423 nm is the least susceptible to the influence of the environmental temperature, if the measurement wavelength is 423 nm or a wavelength close thereto, the environment This is because it is considered that the glycohemoglobin concentration can be measured accurately and stably without being greatly affected by the temperature.

  As the measurement wavelength, as described above, 520-526 nm or 583-589 nm in which the molecular extinction coefficient of oxyhemoglobin and the molecular extinction coefficient of deoxyhemoglobin match or substantially match can be adopted. Is one that selectively transmits light in the wavelength range of 520 to 526 nm or 583 to 589 nm.

  The reference light receiving system 54 shown in FIG. 2 is for acquiring data for suppressing the influence of the turbidity and scattering of the eluent from the analysis column 40, and is reflected by the beam splitter 52 and has an optical path. Of the changed light, the reference wavelength of 500 nm is selectively received. The measurement light receiving system 74 includes an interference filter 54A that selectively transmits light having a wavelength of 500 nm, and a light receiving element 54B that receives light transmitted through the interference filter 54A. A photodiode can be used as the light receiving element 54B.

  As shown in FIG. 4, the arithmetic circuit 6 includes a control unit 60 and a calculation unit 61.

  The control unit 60 is for controlling the operation of each unit. More specifically, the control unit 60 controls turning on / off of the light source 51, controls the interference filter 53A, selects the wavelength of light received by the light receiving element 53B, or performs density calculation processing in the calculation unit 61. Control.

  The calculation unit 61 is for calculating the glycohemoglobin concentration in whole blood based on the light reception results of the light receiving elements 53B and 54B. The calculation unit 61 stores a program necessary for calculation, and its operation is controlled by the control unit 60.

  Next, the operation of the HPLC apparatus X will be described with reference to the flowcharts shown in FIGS. 5 and 6 in addition to FIGS. 1 to 4.

  As shown in FIGS. 1 and 5, in the HPLC apparatus X, when an instruction to start measurement is confirmed (S1), an eluent is supplied to the analysis column 40 (S2). The eluent is supplied from the eluent bottles 10, 11, and 12 to the injection valve 43 through the deaerator 2 and the manifold 41 by the power of the liquid feed pump 42, and the eluents of the plurality of eluent bottles 10, 11, and 12 are also supplied. Which of the eluent bottles 10, 11, and 12 is to be supplied is selected by controlling the manifold 41. The eluent supplied to the injection valve 43 is supplied to the analysis column 40 via the pipe 74.

  In the HPLC apparatus X, a sample for introduction to be introduced into the analytical column 40 is further prepared (S3). In preparing the sample for introduction, a blood sample 14 is first collected from the blood collection tube 13. Collection of the blood sample 14 from the blood collection tube 13 is performed by operating the sampling nozzle 30. The blood sample 14 collected by the sampling nozzle 30 is supplied to the dilution tank 32 by operating the sampling nozzle 30. Further, a hemolyzing agent and a diluent are sequentially supplied from the preparation liquid tank 31 to the dilution tank 32, and a sample for introduction is prepared by mixing the liquid in the dilution tank 32 by pipetting operation using the sampling nozzle 30. .

  The introduction sample is introduced into the analysis column 40 (S4). The introduction sample is introduced into the analysis column 40 by switching the injection valve 43 so that the eluent flows through the injection loop 44. That is, the sample for introduction of the injection valve 44 is introduced into the analysis column 40 together with the eluent. In the analysis column 40, glycohemoglobin is adsorbed to the filler by introducing the introduction sample. After glycated hemoglobin is adsorbed on the filler, the type of eluent supplied to the analysis column 40 is appropriately switched by the manifold 41 to elute the glycated hemoglobin adsorbed on the filler.

  On the other hand, when a certain period of time has elapsed since the introduction of the introduction sample, the eluent is continuously supplied to the analytical column 40 and the injection loop 44 is washed by switching the injection valve 43 ( S5). On the other hand, at the same time as the cleaning of the injection loop 44, a sample for introduction is prepared from the blood sample 14 of the blood collection tube 13 different from the previous one in the same manner as described above (S3). In step S4, the sample for introduction is again introduced into the injection loop 44. Such introduction sample preparation (S3), introduction (S4), and washing (S5) are repeated according to the number of blood collection tubes 13 (blood samples 14) to be measured while appropriately switching the injection valve 43. It is.

  The eluent containing glycohemoglobin discharged from the analysis column 40 is supplied to the photometric cell 50 of the photometric mechanism 5 through the pipe 76 as shown in FIG. 2 and photometrically measured (S6). The eluent from the analysis column 40 is introduced into the photometric cell 50 through the pipe 75 and the introduction flow path 50A, and this eluent passes through the photometric flow path 50B and the discharge flow path 50C, and then from FIG. As can be seen, the liquid is introduced into the waste liquid tank 15 through the pipe 76.

  As shown in FIG. 2, in the photometric mechanism 5, when the eluent from the analysis column 40 passes through the photometric channel 50 </ b> B, the light source 51 continuously irradiates the eluent with light. On the other hand, the light transmitted through the photometric channel 50B is split by the beam splitter 52 and then received by the measurement light receiving system 53 and the reference light receiving system 54. In the measurement light receiving system 53, the light transmitted through the interference filter 53A is selectively received by the light receiving element 53B. On the other hand, in the reference light receiving system 54, light having a reference wavelength of 500 nm transmitted through the interference filter 54A is selectively received by the light receiving element 54B.

  The results of light reception by the light receiving elements 53B and 54B are output to the arithmetic circuit 6, and the arithmetic circuit 6 calculates the concentration of glycohemoglobin (S7). The density calculation processing in the calculation circuit 6 is performed according to the procedure of the flowchart shown in FIG.

  First, the absorbance corresponding to hemoglobin is integrated (S10), and the absorbance corresponding to glycohemoglobin (the portion indicated by cross hatching in FIG. 7) is integrated (S11).

  Next, the ratio of the integrated value of S11 in the integrated value of absorbance in S10 is calculated, and this is used as the glycohemoglobin concentration (S12).

  When the calculation in S12 is completed, the process returns to S7 in FIG. 5 (S13). That is, the calculation result in the calculation circuit 6 is displayed on a display panel (not shown) and printed out automatically or by a user's button operation (S8).

  In the present embodiment, the glycohemoglobin concentration is calculated using the wavelength at which the molecular extinction coefficient of oxyhemoglobin and the molecular extinction coefficient of deoxyhemoglobin match or substantially match as the measurement wavelength. The wavelength at which the molecular extinction coefficients of oxyhemoglobin and deoxyhemoglobin match is constant without being affected by the ratio of oxyhemoglobin and deoxyhemoglobin. Stable measurement is possible regardless of the ratio of oxyhemoglobin and deoxyhemoglobin. As a result, in the present embodiment, when measuring the glycohemoglobin concentration in an environment where the temperature (environment temperature) outside the HPLC apparatus X fluctuates or in a different environment temperature, or the oxygen concentration in the sample introduced into the analytical column Even when this variation occurs, stable glycated hemoglobin concentration can be measured regardless of the ratio of oxyhemoglobin to deoxyhemoglobin in the eluent.

  The present invention is not limited to the embodiment described above, and various modifications can be made. For example, in the concentration calculation process described above, the hemoglobin amount is acquired as the absorbance. However, the hemoglobin amount is not necessarily acquired as the absorbance, and may be acquired as the transmittance or simply as the received light amount. .

  Further, in the photometric mechanism 5, the configuration for causing the light receiving element 53B to receive light having a target wavelength (for example, light having a wavelength range of 417 to 426 nm) is not limited to the configuration using the interference filter 53A, and is publicly known. Other methods can also be adopted.

  The present invention is not limited to an HPLC apparatus for measuring the concentration of glycated hemoglobin in blood, but is also applied to a sample other than blood, or to a liquid chromatography apparatus other than the HPLC apparatus or other glycated hemoglobin concentration measuring apparatus. can do.

  In this example, the influence of the measurement wavelength on the measured value of glycohemoglobin was examined, and the relationship between the environmental temperature and the measurement wavelength was examined.

  Regarding the concentration of glycated hemoglobin, the ambient temperature is set to 10 ° C., 20 ° C., and 30 ° C. In a glycated hemoglobin measuring device (“ADAMS A1c HA-8160”; manufactured by ARKRAY, Inc.), a photodiode array (“ The measurement was performed using a UV-visible multi-wavelength detector MD-910 "(manufactured by JASCO Corporation). The glycated hemoglobin concentration was calculated as the ratio of the glycated hemoglobin amount to the total hemoglobin amount for every 1 nm in the wavelength range of 415 to 430 nm.

  As samples, blood collected from healthy persons (healthy person samples) and blood collected from diabetic patients (diabetic patient blood) were used. The measurement results of glycohemoglobin when using a healthy human specimen are shown in Table 1 and FIG. 8A below, and the measurement results of glycohemoglobin when using a diabetic patient specimen are shown in Table 2 and FIG. 8B below.

  For healthy human samples, as can be seen from Table 1 and FIG. 8A, when the measurement wavelength was 423 nm, the measurement values substantially matched regardless of the environmental temperature. Further, when the measurement wavelength was in the range of 416 to 427 nm, the measurement wavelength was the maximum absorption wavelength of oxyhemoglobin, and it was less affected by the environmental temperature than when it was 415 nm, and the measurement value was stable. In particular, when the measurement wavelength was 419 to 426 nm, it was not further influenced by the environmental temperature.

  On the other hand, as can be seen from Table 2 and FIG. 8B, the measured values of the diabetic patient samples substantially coincided regardless of the environmental temperature when the measurement wavelength was 423 nm. In addition, when the measurement wavelength was in the range of 417 to 427 nm, the measurement wavelength was the maximum absorption wavelength of oxyhemoglobin, and was less affected by the environmental temperature than when it was 415 nm. In particular, when the measurement wavelength was 419 to 425 nm, it was not further influenced by the environmental temperature.

  From the results of this example, when measuring the glycated hemoglobin concentration, if the measurement wavelength is set to 417 to 427 nm, preferably set to 419 to 425 nm, most preferably 423 nm, it is a healthy sample or a diabetic patient. Regardless of whether it is a specimen or not, it can be seen that it is less affected by the environmental temperature and can perform accurate and stable measurement.

  In this example, when the measurement wavelength is set to 415 nm, which is the maximum absorption wavelength of oxyhemoglobin, and when the measurement wavelength is set to 423 nm where the best result is obtained in Example 1, the environmental temperature becomes the measurement value. The effect was examined.

  The measured value of glycated hemoglobin was measured for each of the healthy subject sample and the diabetic patient sample in the same manner as in Example 1. The measurement results of glycohemoglobin when the measurement wavelength is set to 415 nm are shown in Table 3 and FIG. 9A below, and the measurement results when the measurement wavelength is set to 423 nm are shown in Table 4 and FIG. 9B below.

  As shown in Table 3 and FIG. 9A, when measuring glycated hemoglobin with the measurement wavelength set to 415 nm, which is the maximum absorption wavelength of oxyhemoglobin, the measured value increases as the environmental temperature increases, and the measured value becomes It was greatly affected by temperature.

  On the other hand, as shown in Table 4 and FIG. 9B, when the glycohemoglobin concentration was calculated with the measurement wavelength set to 423 nm, the measurement was performed even if the environmental temperature changed in the range of 10 to 30 ° C. The value was not significantly affected by the environmental temperature, and became a substantially constant value. In particular, in a diabetic patient specimen that requires more accurate measurement, there was almost no variation in the measured value with respect to the environmental temperature. For this reason, when the glycohemoglobin concentration is calculated with the measurement wavelength set to 423 nm, accurate and stable glycolysis is not affected by the environmental temperature (dissolved oxygen concentration of the eluent) or the dissolved oxygen concentration of the sample. It can be seen that the hemoglobin concentration can be measured. Further, even when the measurement wavelength is set to a wavelength close to 423 nm, it is presumed that the accurate and stable measurement of glycated hemoglobin can be performed without being greatly affected by the environmental temperature.

  From the above, it was found that by selecting a wavelength close to 423 nm, the concentration of glycated hemoglobin can be accurately measured regardless of the ratio of oxyhemoglobin and deoxyhemoglobin.

Claims (12)

A method for measuring glycated hemoglobin concentration by an optical method,
A glycohemoglobin concentration measuring method, wherein a measurement wavelength is set to a wavelength at which molecular extinction coefficients of oxyhemoglobin and deoxyhemoglobin match or substantially match.   The glycohemoglobin concentration measuring method according to claim 1, wherein the measurement wavelength is set to 417 to 427 nm.   The glycohemoglobin concentration measuring method according to claim 1, wherein the measurement wavelength is set to 419 to 425 nm.   The method for measuring the concentration of glycohemoglobin according to claim 1, wherein the measurement wavelength is set to 520 to 526 nm or 583 to 589 nm.   The method for measuring glycated hemoglobin concentration according to claim 1, wherein the glycated hemoglobin concentration is measured using column chromatography.   The method for measuring glycated hemoglobin concentration according to claim 1, wherein the glycated hemoglobin concentration is measured using a sample prepared from erythrocytes in blood. A glycated hemoglobin concentration measuring device equipped with a photometric mechanism that irradiates a sample with light from a light source and receives light traveling from the sample at that time in a light receiving unit,
The glycated hemoglobin concentration measuring apparatus, wherein the photometric mechanism is configured such that light having a wavelength at which molecular extinction coefficients of oxyhemoglobin and deoxyhemoglobin match or substantially match can be received by a light receiving unit.   The glycated hemoglobin concentration measuring apparatus according to claim 7, wherein the photometric mechanism is configured to receive light of 417 to 427 nm at the light receiving unit.   The glycated hemoglobin concentration measuring apparatus according to claim 7, wherein the photometric mechanism is configured so that light of 419 to 425 nm can be received by the light receiving unit.   The glycated hemoglobin concentration measuring apparatus according to claim 7, wherein the photometric mechanism is configured so that light of 520 to 526 nm or 583 to 589 nm can be received by the light receiving unit.   The glycated hemoglobin concentration measuring device according to claim 7, further comprising a separation unit for separating glycated hemoglobin using column chromatography.   The glycated hemoglobin concentration measuring apparatus according to claim 7, further comprising a sample preparation mechanism for preparing a sample from red blood cells in blood.

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